In this project we propose to follow up on a number of observations that we have made concerning CD4+ T cells and how they may contribute to immune competence, especially in young children. Recently we found that healthy adults have an abundance of memory phenotype CD4+ T cells (which we refer to as TMPS) that are specific for viral antigens to which they have never been exposed. This is in contrast to newborns, where we find a similar frequency of specific T cells, but almost entirely naive in phenotype. These TMPS in adults have many of the expected characteristics of authentic memory T cells, and we hypothesize that they are the consequence of microbial exposure to cross-reactive T cell receptors (TCR), confer some degree of immunological protection, and contribute to immunodominance and a fully capable immune system. In Aim 1, we will characterize the time course with which specific TMRS appear in infants and children and also analyze whether their rise is continuous, or spikes with major vaccinations, acquisition of a microbiome or disease exposure. For the purposes of comparison to children recruited here, we will also have age and gender matched samples from our collaborators in Bangladesh. In Aim 2, we will perform a critical test of whether and to what degree cells of this type participate in a vaccine response, by first analyzing the TCR repertoires of naive and TMRS specific for the same Hepatitis B antigens and then seeing which TCRs in the same subject are expanded in the effector and then memory T pools after challenge with the vaccine. We will also use this same procedure to analyze the response to unique flu strain epitopes. Another effort will be to find original cross-reactive epitopes for selected TMRS using unique peptide-MHC display libraries, which we have constructed. Lastly, in Aim 3, we wish to follow up on our studies of flu specific effector and memory CD+ T cells, where we see evidence of an influence on both antibody responses and specific CD8+ T cell levels. The most prominent of these T cells responding to flu vaccination have some of the characteristics of Follicular helper T cells and may be driving the responses of the other cells. Using the novel ex vivo infection system described in the Greenberg project (P2) will afford us a way to directly test this hypothesis.